Myocardial infarction is prevalent with -1 million patients diagnosed each year. To better understand the cellular conditions leading to cardiomyocyte damage and death, experimental models have mimicked the ischemia - reperfusion (I-R) experienced by the myocardium. The decrease in ATP with I-R is often considered the driving force behind the contractility decline. However, recent research suggests that changes intrinsic to the contractile filaments, such as protein proteolysis or redox-dependent protein modifications, also influence contractility during I-R. The preliminary data in this application indicate that a decline in cardiac muscle contractility occurs with 30' of ischemia and is largely reversed by 60' of reperfusion. The reversible decline in contractility was independent of ATP availability, suggesting that intrinsic changes to the contractile filaments best described the decline. However, this timeframe is insufficient for protein proteolysis during ischemia to be rescued by protein synthesis and re-assembly during reperfusion. Therefore, these changes in contractility may reflect reversible, covalent modifications to proteins of the contractile filaments rather than their proteolysis. Consistent with this hypothesis, preliminary data demonstrate that a reversible modification of actin occurs during I-R, affecting it's interaction with tropomyosin. Therefore, this grant application aims to investigate fibre contractility during I-R, and characterize the reversible modifications to proteins of the contractile filaments that underlie the changes in contractility. The application will test the hypothesis that ischemia-reperfusion results in reversible, covalent modifications to proteins of the cardiac muscle thin filament, consequently limiting contractility through changes in the association of thin filament regulatory proteins. This hypothesis will be examined by: i) determining the effect of I-R on the contractility of cardiac muscle fibres; ii) characterizing the I-R dependent modification of actin, and determining if I-R results in covalent modifications to other thin filament proteins; iii) determining the effect of modification of actin on thin filament assembly as well as the actin activated myosin ATPase. These findings will provide novel insight into the nature of the contractile deficit during I-R, with emphasis on the state of the cardiac muscle thin filament proteins and their effect on contractility.